Brucella abortus (hereinafter "B. abortus") is a bacterial organism which causes brucellosis which is characterized by spontaneous abortion and chronic infection within the lymph nodes and in the mammary glands of animals such as cattle or bison. This disease causes extensive economic loss due to abortion, the birth of weak debilitated calves, decreased milk production and infertility.
To reduce the incidence and economic loss caused by brucellosis, animals are often vaccinated with either live or killed organisms of B. abortus. Production of antibodies specific for B. abortus is a serological defense against bovine brucellosis and is the partial premise for prophylactic vaccination. The vaccine of choice is the attenuated B. abortus strain 19, which like the field Strain 2308, is strongly antigenic. Unfortunately, a consistently effective means for distinguishing Strain 19 vaccinated animals from those infected by pathogenic strains of B. abortus or Strain 19 is not available. Both the vaccine and the pathogenic strains stimulate the production of cross-reacting antibodies which serve as the basis for current serological tests for brucellosis. Hence, the current serological tests are often unable to distinguish Strain 19 vaccinated animals from field strain or Strain 19 infected animals, including cattle and bison. Inaccurate diagnosis often results in economic losses due to unnecessary testing of herds and unnecessary quarantine of cattle which in turn inhibits the movement and marketing of cattle through livestock marketing systems. Thus, a need exists for an assay which is capable of distinguishing between infected and vaccinated animals.
The current procedures used to detect antibodies stimulated by B. abortus include the card test, rivanol, particle concentration fluorescence immunoassay (PCFIA), hemolysis-in-gel (HIG), complement fixation (CF) and the indirect enzyme-linked immunosorbant assay (ELSIA). However, none of these current tests have proven to be effective in differentially detecting Strain 19 stimulated antibodies versus field strain stimulated antibodies.
The antibodies detected by the CF and ELISA tests correlate closely with the presence of disease. Unfortunately, even these tests can not detect all infected cattle without giving false positive reactions in uninfected cattle. Hence, in practice, the serological testing component of a brucellosis eradication program often consists of a combination of serological tests. The inefficiency and often contradictory results of these combined procedures often affect the progress toward eradication of the disease.
For diagnostic usefulness, a definitive test must have a high degree of accuracy in successfully identifying both:
(1) infected animals in a diseased population (high sensitivity); and PA1 (2) disease-free individuals in a healthy population (high specificity).
The sensitivities reported for the current CF and ELISA procedures range from 86-93% and 87-89% respectively. Similarly, the specificities reported for these two tests respectively are 93-99% and 26-51%.
Recently, two ELISA procedures have been proposed to differentiate between vaccinated and infected cattle. More specifically, a first ELISA assay system for differentially detecting antibodies stimulated by Strain 19 vs. field strains utilized a monoclonal antibody to Yersinia enterocoIitica 0:9 smooth lipopolysaccharide antigen. However, this assay lacked sufficient sensitivity because it failed to detect cattle infected with low doses of field Strain 2308 of B. abortus (see Nielsen et al., Am. J. Vet. Res., Vol 50, No. 1, (1989) 5-9). A second ELISA procedure utilized a monoclonal antibody to B. abortus 544 smooth lipopolysaccharide antigen. However, this assay lacked sufficient specificity as it failed to differentiate at least 10% of vaccinated cattle from infected cattle. Also, the assay is based upon a cumbersome ELISA procedure which offers a limited possibility for mass-scale production (see Chin et al., Vet. Immunol. Immunopathol., Vol. 20, (1989) 109-118).